Antiphospholipid antibodies (aPA) are a heterogeneous group of circulating autoantibodies that were originally thought to bind negatively-charged phospholipids. These autoantibodies have been associated with a range of clinical associations, in particular thrombotic events. Despite extensive literature in the pathogenic mechanisms, which explain the association between aPA and the major clinical manifestations of the antiphospholipid syndrome, mainly thrombosis, are unknown. These autoantibodies were originally thought to bind negatively-charged phospholipids and could be demonstrated in a variety of other clinical syndromes. However, in 1990 the present inventor demonstrated for the first time that these autoantibodies in fact bind β2-glycoprotein I (β2-GPI), a phospholipid binding plasma protein whose physiological role is unknown.
Currently there are two assay systems in clinical practice to identify these autoantibodies. The first assay uses the negatively-charged phospholipid most commonly cardiolipin to coat microtitre wells and antibodies detected in this assay are known as anticardiolipin (ACL) antibodies. This assay system detects anticardiolipin antibodies in autoimmune patients and also in a variety of infections which are not associated with thrombotic events. It appears now that the autoimmune ACL antibodies detected in the ELISA system are directed against β2-GPI, which is captured on the negatively charged surface. The anticardiolipin antibodies occurring in infections like malaria, syphilis, leprosy, tuberculosis are not associated with clinical thrombotic events are now known not to bind β2-GPI but in fact are directed against negatively-charged phospholipids and are thus true phospholipid antibodies. These antibodies are usually of low affinity and are dependent on charge as high salt concentrations eliminate their binding to cardiolipin.
The second type of assay used is one that looks at the prolongation of in-vitro clotting assays. Autoantibodies that prolong phospholipid-dependent clotting tests which are not corrected by adding normal plasma and are neutralised by the addition of phospholipid are defined as lupus anticoagulants (LA). Patients may have either of these antibody subtypes or both activities. More recently the use of β2-GPI in an ELISA identifies the autoimmune ACL antibodies since the infective ACL antibodies do not bind in this assay system. However, for clinical testing, both assays have to be utilised as patients with the antiphospholipid syndrome may be missed if only one assay system is used.
The present inventor has now developed an in-vitro assay system which is simple, will identify both ACL and LA type antibodies, would discriminate between infective and autoimmune type ACL autoantibodies and may be used to identify patients at risk of clinical thrombotic events.